Reverse cholesterol transport (RCT) is a term used to describe the efflux of excess cellular cholesterol from peripheral tissues and its return to the liver for excretion in the bile and ultimately the feces. It is believed to be a critical mechanism by which HDL exert a protective effect on the development of atherosclerosis. In this paradigm, cholesterol is effluxed from arterial macrophages to extracellular HDLbased acceptors through the action of transporters such as ABCA1 and ABCG1. After efflux to HDL, cholesterol may be esterified in the plasma by the enzyme lecithin:cholesterol acyltransferase and is ultimately transported from HDL to the liver, either directly via the scavenger receptor BI or after transfer to apolipoprotein B-containing lipoproteins by the cholesteryl ester transfer protein. Methods for assessing the integrated rate of macrophage RCT in animals have provided insights into the molecular regulation of the process and suggest that the dynamic rate of macrophage RCT is more strongly associated with atherosclerosis than the steady-state plasma concentration of HDL cholesterol. Promotion of macrophage RCT is a potential therapeutic approach to preventing or regressing atherosclerotic vascular disease, but robust measures of RCT in humans will be needed in order to confidently advance RCT-promoting therapies in clinical development.-

Genome-wide association studies (GWAS) have identified a genetic variant at a locus on chromosome 1p13that is associated with reduced risk of myocardial infarction, reduced plasma levels of LDL cholesterol (LDL-C), and markedly increased expression of the gene sortilin-1 (SORT1) in liver. Sortilin is a lysosomal sorting protein that binds ligands both in the Golgi apparatus and at the plasma membrane and traffics them to the lysosome. We previously reported that increased hepatic sortilin expression in mice reduced plasma LDL-C levels. Here we show that increased hepatic sortilin not only reduced hepatic apolipoprotein B (APOB) secretion, but also increased LDL catabolism, and that both effects were dependent on intact lysosomal targeting. Loss-of-function studies demonstrated that sortilin serves as a bona fide receptor for LDL in vivo in mice. Our data are consistent with a model in which increased hepatic sortilin binds intracellular APOB-containing particles in the Golgi apparatus as well as extracellular LDL at the plasma membrane and traffics them to the lysosome for degradation. We thus provide functional evidence that genetically increased hepatic sortilin expression both reduces hepatic APOB secretion and increases LDL catabolism, providing dual mechanisms for the very strong association between increased hepatic sortilin expression and reduced plasma LDL-C levels in humans.

The structure of apoA-I on discoidal high density lipoprotein (HDL) was studied using a combination of chemical cross-linking and mass spectrometry. Recombinant HDL particles containing 145 molecules of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine and two molecules of apoA-I with a 96-Å diameter were treated with the lysine-specific cross-linker, dithiobis(succinimidylpropionate) at varying molar ratios from 2:1 to 200:1. At low molar ratios of dithiobis(succinimidylpropionate) to apoA-I, two products were obtained corresponding to ϳ53 and ϳ80 kDa. At high molar ratios, these two products merged, yielding a product of ϳ59 kDa, close to the theoretical molecular mass of dimeric apoA-I. To identify the intermolecular cross-links giving rise to the two different sized products, bands were excised from the gel, digested with trypsin, and then analyzed by liquid chromatography-electrospray-tandem mass spectrometry. In addition, tandem mass spectrometry of unique cross-links found in the 53-and 80-kDa products suggested that a distinct conformation exists for lipid-bound apoA-I on 96-Å recombinant HDL, emphasizing the inherent flexibility and malleability of the N termini and its interaction with its C-terminal domain.The structure of apolipoprotein A-I (apoA-I) 2 has been intensely investigated in efforts to understand its highly significant role in protecting against coronary heart disease in humans (1-3). ApoA-I is abundantly found in plasma high density lipoproteins (HDL) and functions as the main carrier of excess cholesterol to the liver in a process termed "reverse cholesterol transport" (4 -6). ApoA-I also plays a significant role in mediating anti-inflammatory/antioxidative processes (7-12) intervening in the escalation of damage to the artery wall. As with many proteins, the functional roles played by apoA-I are tightly coupled to the structure of the apoprotein. Thus, the lack of a detailed lipid-bound apoA-I x-ray crystal structure has seriously hindered our understanding of this apoprotein's unique features (13,14) in its biologically active lipid-bound form. A major advance occurred when the x-ray crystal structure of lipid-free ⌬43 apoA-I was reported (15). This report suggested that apoA-I adopts an antiparallel "belt-like" conformation when bound to a lipid surface. Unfortunately, lipid-bound apoA-I has not yielded crystals of the quality needed to solve its three-dimensional conformation. Instead, a number of alternative and highly innovative approaches have been used to probe the conformation suggested by the lipid-free crystal structure (16 -27). In addition, computer modeling studies suggest that the two molecules of apoA-I wrap in an extended belt completely around the edge of a lipid bilayer, maximizing intermolecular salt bridges, which act to stabilize the protein conformation in an antiparallel arrangement (23). Although all of these studies support the concept of a "belt" model of apoA-I, their inability to distinguish between an "extended belt" or a "hairpin" belt conformation have lead...